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Kim Nguyen-Phuoc, Gavin Neil, Tim Footz, Adrian Lahola-Chomiak, Ordan J Lehmann, Michael A Walter, W. Ted Allison; Dissecting the Role of the Premelanosome Protein Gene (PMEL) in the Development of Pigmentary Glaucoma Using the Zebrafish Animal Model. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5156.
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© ARVO (1962-2015); The Authors (2016-present)
Premelanosome Protein (PMEL) has been identified as a candidate gene for the development of Pigment Dispersion Syndrome (PDS) and Pigmentary Glaucoma (PG) in humans. Mutations in PMEL have previously been shown to cause pigment and ocular defects in several animals but there is currently no known PMEL associated defects in humans. To better understand the pathology of PDS/PG, we are analyzing the impact that PDS/PG associated variants in PMEL have on the structure and function of the zebrafish eye. Zebrafish are an excellent animal model for ophthalmology because of their shared structure to the human retina developed for day vision and its genetic tractability. Introducing PDS/PG patient mutations in PMEL into zebrafish using transgenics can elucidate the role PMEL variants play in the development of PDS/PG in humans.
We deployed morpholinos (MOs) and CRISPR/Cas9 to generate transient pmela knockdowns and pmela knockout zebrafish, respectively, to assess the requirement for PMEL in early development and ocular maintenance. We assayed Ocular pigmentation, anterior segment structure, retinal health, retinal ganglion cell morphology, and optomotor response .
MOs targeted at a PMEL paralog in zebrafish (pmela) significantly reduced pigmentation compared to embryos injected with a standard control morpholino at 2 days post fertilization (dpf) and 3 dpf in the eyes (p<0.001 for 2 dpf and 3 dpf) and the body (p<0.001 for 3dpf). This phenotype is similar to the zebrafish PMEL mutant, fading vision, which harbors a premature stop codon in the RPT domain of pmela. Additionally, we have successfully used CRISPR/Cas9 to disrupt pmela so as to disrupt the C-terminus of the pmela protein, resulting in several observable ocular defects.
We are assessing PMEL associated pathogenicity by reintroducing PDS/PG associated PMEL variants into a vertebrate model. Using zebrafish as an animal model to ascertain the role of PMEL in the etiology of PDS/PG will help elucidate the mechanisms of the disease, leading to novel diagnosis and treatment avenues.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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